Factor VII is a precursor to a serine protease that is present in mammalian plasma. In its activated form, it participates in blood coagulation by activating factor X and/or factor IX in the presence of tissue factor and calcium. Clones coding for factor VII were obtained from two cDNA libraries prepared from poly(A) RNA from human liver and Hep G2 cells. The amino acid sequence deduced from the cDNAs indicates that factor VII is synthesized with a prepro-leader sequence of 60 or 38 amino acids. The mature protein that circulates in plasma is a single-chain polypeptide composed of 406 amino acids. The amino acid sequence analysis of the protein and the amino acid sequence deduced from the cDNAs indicate that factor VII is converted to factor VII8 by the cleavage of a single internal bond between arginine and isoleucine. This results in the formation of a light chain (152 amino acids) and a heavy chain (254 amino acids) that are held together by a disulfide bond. The light chain contains a -carboxyglutamic acid (Gla) domain and two potential epidermal growth factor domains, while the heavy chain contains the serine protease portion of the molecule. Factor VII shows a high degree of amino acid sequence homology with the other vitamin K-dependent plasma proteins.

Infestation of the gastrointestinal tract by parasitic nematodes is invariably associated with mucosal mastocytosis, which is a thymus-dependent phenomenon in parasitized rats, and is adoptively transferable with a T cell-enriched population of thoracic duct lymphocytes. When derived by in vitro culture, mucosal mast cells (MMC) arise from a bone marrow precursor after stimulation by T cell-derived factors. In rats infected with the nematode Trichinella spiralis, mucosal mastocytosis is temporally associated with the immune expulsion of the adult worms whereas in the case of Nippostrongylus brasiliensis, mastocytosis is frequently observed to occur after worm expulsion has been completed. Consequently, there has been doubt as to whether MMC are active and serve a functional role in the expulsion of rat intestinal nematodes. MMC contain and secrete a neutral proteinase, rat mast cell protease II (RMCP II); detection and assay of secreted RMCP II therefore provides a direct measurement of MMC activity. Here we describe the release of this enzyme into the blood of rats infected with N. brasiliensis or T. spiralis. Our results show that the systemic secretion of RMCP II coincides with the immune expulsion of these nematodes, demonstrating clearly for the first time that rat MMC are functionally active during the immune elimination of primary nematode infections.

Secretory granules of the rat basophilic leukemia (RBL-1) cell, a chemically-generated tumor cell line maintained in tissue culture, were shown to stain with alcian blue but not with safranin counterstain and to have sparse, small, electron-dense granules. A Mr 25,000 protein was the major [3H]diisopropyl fluorophosphate-binding protein in extracts of RBL-1 cells. Double-immunodiffusion analysis of extracts revealed immunoreactivity for rat mast cell protease (RMCP)-II, a Mr 25,000 neutral protase present in the secretory granules of rat mucosal mast cells and cultured rat bone marrow-derived mast cells, but no immunoreactivity for RMCP-I, the predominant neutral protease of rat connective tissue mast cells. By radial immunodiffusion, there was 66.8 ng of RMCP-ll per 106 cells. Whereas rat connective tissue mast cells stain with alcian blue and safranin and contain heparm proteoglycan, rat mucosal and rat bone marrow-derived mast cells stain with alcian blue only and contain a non-heparin proteoglycan and lesser amounts of histamine. Proliferation of rat mucosal mast cells in vivo and rat bone marrowderived mast cells in vitro requires T-cell factors, whereas no comparable requirement has been observed for connective tissue mast cells. The transformed RBL-1 tumor cell, whose growth is independent of factors other than those present in standard tissue culture medium, has previously been shown to contain predominantly chondroitin sulfate di-B proteoglycans and low amounts of histamine. The similar histology and secretory granule biochemistry of the rat mucosal mast cell, rat culture-derived mast cell, and RBL-1 cell suggest that they comprise a single mast cell subclass distinct from the rat connective tissue mast cell. (6)(7)(8)(9). Both of these rat mast cell subclasses can be activated with IgE and specific antigen. The connective tissue cell also responds to compound 48/80, (a condensation product of N-methyl-pmethoxyphenethylamine and formaldehyde), whereas the mucosal mast cell in situ (10) or isolated from intestine (11) does not. Disodium cromoglycate and theophylline inhibit the activation of the connective tissue mast cell but not of the mucosal mast cell (12). Homogeneous populations of mast cells resembling the mucosal mast cell subclass have been obtained in vitro from rat bone marrow cultured in the presence of conditioned medium from antigen-activated immune mesenteric lymph nodes (13,14). These mast cells stain with alcian blue but not safranin, have low amounts of histamine, contain RMCP-II, and appear to have a non-heparin proteoglycan (15).Thymic-dependent proliferation of mast cells also occurs in the mucosa of helminth-infected mice (16), and mouse hematopoietic stem cells cultured in vitro in the presence of lymphocyte conditioned medium differentiate into mast cells resembling the mucosal mast cell subclass (17-22). The cultured cells depend on the T-cell lymphokine interleukin 3 for growth (23) and contain an oversulfated chondroitin sulfate proteoglycan, termed chondroitin sulfate...

We have used two highly sensitive assays to quantitate p97, a protein associated with human melanoma, in cultured cells and normal adult, fetal, and neoplastic tissues. To measure p97 at the surface of intact cells, radiolabeled Fab fragments of a monoclonal antibody specific for p97 were used in a binding assay. To measure p97 in detergent-solubilized membrane preparations, we used a novel double-determinant immunoassay that uses two monoclonal antibodies to two distinct antigenic determinants of p97. These assays revealed that although p97 is present in small amounts in normal adult tissues, it is present in much larger amounts in most melanomas, in some other tumors (both benign and malignant), and in certain fetal tissues. We conclude that monoclonal antibodies to p97 may prove to be of value for the diagnosis and therapy ofmelanoma.